Bottom Line:
Two of them, chi18-13 and chi18-17, are members of the B1/B2 chitinase subgroup that have expanded significantly in paralog number in mycoparasitic Hypocrea atroviridis and H. virens.Differences in amino acid diversity/conservation patterns between different Trichoderma clades are observed.These observations show that Trichoderma chitinases chi18-13 and chi18-15 evolve in a manner consistent with rapid co-evolutionary interactions and identifies putative target regions involved in determining substrate-specificity.

ABSTRACTCertain species of the fungal genus Trichoderma are potent mycoparasites and are used for biological control of fungal diseases on agricultural crops. In Trichoderma, whole-genome sequencing reveal between 20 and 36 different genes encoding chitinases, hydrolytic enzymes that are involved in the mycoparasitic attack. Sequences of Trichoderma chitinase genes chi18-5, chi18-13, chi18-15 and chi18-17, which all exhibit specific expression during mycoparasitism-related conditions, were determined from up to 13 different taxa and studied with regard to their evolutionary patterns. Two of them, chi18-13 and chi18-17, are members of the B1/B2 chitinase subgroup that have expanded significantly in paralog number in mycoparasitic Hypocrea atroviridis and H. virens. Chi18-13 contains two codons that evolve under positive selection and seven groups of co-evolving sites. Chi18-15 displays a unique codon-usage and contains five codons that evolve under positive selection and three groups of co-evolving sites. Regions of high amino acid variability are preferentially localized to substrate- or product side of the catalytic clefts. Differences in amino acid diversity/conservation patterns between different Trichoderma clades are observed. These observations show that Trichoderma chitinases chi18-13 and chi18-15 evolve in a manner consistent with rapid co-evolutionary interactions and identifies putative target regions involved in determining substrate-specificity.

Mentions:
Amplification products and partial sequences for chi18-15 orthologs, presumably lacking 20–29 amino acid residues in the C-terminal part, were obtained from nine different Trichoderma species (Fig. S3). Additional full-length sequences from H. jecorina, H. atroviridis and H. virens were retrieved from the genome sequences. A phylogenetic analysis confirmed the orthologous status of the sequenced genes (Fig. S3). Amino acid diversity was distributed amongst eight regions with W mean scores above the defined threshold (Fig. 4A). Additional analyses of sequences from taxonomic subgroups (see below) identified an additional region of high amino acid diversity in the C-terminal end of chi18-15 (Fig. 4B). Two of these regions were associated with the secretion signal peptide while the other seven regions (Ic, IIc, IIIc, IVc, Vc, VIc and VIIc) were located in the catalytic module shown in Figure 2C prepared using the homology model of H. jecorina chi18-15. Although several of these regions were surface-exposed in the homology model, none of the parts contribute to the catalytic cleft. The seven residues predicted as important for catalysis in chi18-15 (cd02871 in CDD) were all located in conserved regions with low W scores, as were all predicted substrate-interacting residues (Fig. 4A).

Mentions:
Amplification products and partial sequences for chi18-15 orthologs, presumably lacking 20–29 amino acid residues in the C-terminal part, were obtained from nine different Trichoderma species (Fig. S3). Additional full-length sequences from H. jecorina, H. atroviridis and H. virens were retrieved from the genome sequences. A phylogenetic analysis confirmed the orthologous status of the sequenced genes (Fig. S3). Amino acid diversity was distributed amongst eight regions with W mean scores above the defined threshold (Fig. 4A). Additional analyses of sequences from taxonomic subgroups (see below) identified an additional region of high amino acid diversity in the C-terminal end of chi18-15 (Fig. 4B). Two of these regions were associated with the secretion signal peptide while the other seven regions (Ic, IIc, IIIc, IVc, Vc, VIc and VIIc) were located in the catalytic module shown in Figure 2C prepared using the homology model of H. jecorina chi18-15. Although several of these regions were surface-exposed in the homology model, none of the parts contribute to the catalytic cleft. The seven residues predicted as important for catalysis in chi18-15 (cd02871 in CDD) were all located in conserved regions with low W scores, as were all predicted substrate-interacting residues (Fig. 4A).

Bottom Line:
Two of them, chi18-13 and chi18-17, are members of the B1/B2 chitinase subgroup that have expanded significantly in paralog number in mycoparasitic Hypocrea atroviridis and H. virens.Differences in amino acid diversity/conservation patterns between different Trichoderma clades are observed.These observations show that Trichoderma chitinases chi18-13 and chi18-15 evolve in a manner consistent with rapid co-evolutionary interactions and identifies putative target regions involved in determining substrate-specificity.

ABSTRACTCertain species of the fungal genus Trichoderma are potent mycoparasites and are used for biological control of fungal diseases on agricultural crops. In Trichoderma, whole-genome sequencing reveal between 20 and 36 different genes encoding chitinases, hydrolytic enzymes that are involved in the mycoparasitic attack. Sequences of Trichoderma chitinase genes chi18-5, chi18-13, chi18-15 and chi18-17, which all exhibit specific expression during mycoparasitism-related conditions, were determined from up to 13 different taxa and studied with regard to their evolutionary patterns. Two of them, chi18-13 and chi18-17, are members of the B1/B2 chitinase subgroup that have expanded significantly in paralog number in mycoparasitic Hypocrea atroviridis and H. virens. Chi18-13 contains two codons that evolve under positive selection and seven groups of co-evolving sites. Chi18-15 displays a unique codon-usage and contains five codons that evolve under positive selection and three groups of co-evolving sites. Regions of high amino acid variability are preferentially localized to substrate- or product side of the catalytic clefts. Differences in amino acid diversity/conservation patterns between different Trichoderma clades are observed. These observations show that Trichoderma chitinases chi18-13 and chi18-15 evolve in a manner consistent with rapid co-evolutionary interactions and identifies putative target regions involved in determining substrate-specificity.